Determination of Perampanel in Dried Plasma Spots: Applicability to Therapeutic Drug Monitoring.

BACKGROUND: Although therapeutic drug monitoring of antiepileptic drugs is typically based on the analysis of plasma samples, alternative matrices, such as dried plasma spots (DPSs), may offer specific advantages. The aims of this work were to (1) develop and validate a bioanalytical method for the quantitative determination of the second-generation antiepileptic drug perampanel in DPSs; (2) assess short- and long-term stability of perampanel in DPSs; and (3) test the clinical applicability of the developed method. METHODS: Two hundred microliters of plasma were dispensed on a glass paper filter and dried. Glass paper filter discs were then inserted into clean tubes. After addition of the internal standard (ie, promethazine), the analytes were extracted with 5-mL methanol, dried at room temperature (23 ± 2°C), and reconstituted. Separation and quantification were achieved on 2 serial reverse-phase monolithic columns connected to an UV detector (λ = 320 nm). RESULTS: Calibration curves were linear in the validated concentration range (25-1000 ng/mL). Intraday and interday accuracy were in the range of 99.2%-111.4%, whereas intraday and interday precision (coefficient of variation) ranged from 2.8% to 8.6%. The lowest limit of quantitation was 25 ng/mL. The stability of the analyte in DPSs was assessed and confirmed under different storage conditions. Perampanel concentrations estimated in DPS samples from patients receiving therapeutic doses were equivalent to those measured in plasma samples. CONCLUSIONS: This simple method enables the quantitation of perampanel in DPSs with adequate accuracy, precision, specificity, and sensitivity. The short- and long-term stabilities of perampanel in DPSs are highly beneficial for sample shipment or storage at ambient temperature. Moreover, DPSs decreases the costs associated with storage and transportation compared with conventional wet samples.

A simple and rapid HPLC-UV method for the determination of retigabine in human plasma.

A simple and rapid high-performance liquid chromatographic method with ultraviolet detection was developed for the quantitative determination of retigabine, known also as ezogabine, in human plasma. The assay uses a simple solid-phase extraction for sample preparation and direct injection of the extract into the chromatograph. Flupirtine is used as an internal standard. Chromatographic separation is achieved on a C18 Chromolith column (Chromolith Performance, 100 × 4.6 mm i.d.), using as mobile phase water/acetonitrile/methanol (72:18:10 v/v/v) mixed with 0.1% of 85% phosphoric acid. Isocratic elution is conducted at a flow rate of 1.5 mL min-1 . The total duration of a chromatographic run is 7 min. Calibration curves are linear over the 25-2000 ng mL-1 concentration range, with a limit of quantitation of 25 ng mL-1 . Other performance characteristics include high precision (intra- and inter-day coefficients of variation ≤12.6%) and high accuracy (99.7%-108.7%). The method is suitable for the investigation of concentration-response relationships in patients receiving therapeutic doses of retigabine.

A UPLC-MS-MS method for the simultaneous quantification of first-line antituberculars in plasma and in PBMCs.

OBJECTIVES: TB is currently the second cause of death among patients affected with infectious diseases. Quantification of drug levels in plasma and in cells where Mycobacterium tuberculosis persists and grows may be useful in understanding the appropriateness of dosage regimens. We report a new and fully validated chromatographic method to quantify first-line antituberculars in plasma and PBMCs. The method was used for plasma and cell quantification of antituberculars in patients undergoing treatment with standard oral therapy. METHODS: Ethambutol, isoniazid, pyrazinamide and rifampicin were extracted from plasma and PBMCs using two separate and optimized procedures; analysis was performed using UPLC coupled with a mass-mass detector system (UPLC-MS-MS). Antitubercular levels in patients were assayed at the end of the dosing interval (C trough) and 2 h post-dose (C max). RESULTS: The method was accurate and precise. Recovery and the matrix effect were reproducible. While rifampicin intracellular concentrations were similar to plasma values (median intra-PBMC C max = 7503 ng/mL versus median plasma C max = 6505 ng/mL), isoniazid and pyrazinamide intracellular concentrations were lower than plasma values (median intra-PBMC C max = 12 ng/mL versus median plasma C max = 3258 ng/mL for isoniazid and median intra-PBMC C max = 2364 ng/mL versus median plasma C max = 26 988 ng/mL for pyrazinamide) and ethambutol intracellular concentrations were significantly higher than plasma values (median intra-PBMC C max = 73 334 ng/mL versus median plasma C max = 2244 ng/mL). CONCLUSIONS: The method was suitable for both therapeutic drug monitoring and for pharmacokinetic analysis. Should the clinical usefulness of measuring antitubercular drug intracellular concentrations be confirmed, this method could be useful to enhance the clinical application of intra-PBMC evaluation.